HIV-1 specific CD8 cytotoxic T lymphocyte (CTL) responses play a critical role in controlling HIV-1 replication. TCR avidity correlates with CTL function, and CTLs expressing TCRs with high avidity for their cognate MHC-viral peptide complex play an important in vivo role in neutralizing virus infections, terminating virus infection and delaying systemic AIDS virus dissemination from the mucosal inoculation site. Little is known about the structural basis by which the TCR's expressed by HIV-specific CTL recognize the HIV peptide:MHC Class I complex and the contributions of specific TCR residues to the TCR avidity for its cognate HIV peptide. Elucidation of the structure- function correlates of HIV peptide recognition by the TCR expressed by HIV-specific CD8+ CTLs will increase our understanding of the mechanism by which some TCRs confer CTLs with potent anti-HIV activity. We hypothesize that after solving the TCR structure and identifying TCR-peptide contact residues, the functional activity of HIV- specific CTLs can be markedly enhanced by mutation of TCR amino acids that contact the cognate peptide thereby increasing its affinity for its cognate peptide:MHC complex. We have cloned the TCR genes from an HIV-specific CTL clone into lentiviral vectors that efficiently transform peripheral CD8 T cells into HIV-specific CTLs with defined specificities. We have also cloned these TCR genes as a linked single chain TCR (scTCR) into bacterial, drosophila and yeast vectors to express sufficient protein to generate crystals to determine the TCR structure. The structural basis of TCR avidity for its cognate HIV peptide will be obtained by determining the X-ray crystallographic structures of the complexes formed by HLA-A*0201-restricted HIV-SL9-specific TCRs and their cognate MHC Class I:SL9-peptide assemblies. These structures will permit us to 1) define the physical and chemical determinants responsible for MHC:peptide recognition;2) determine/rationalize the effect of targeted mutations on CTL function and 3) perform structure-assisted design of novel TCRs with unique binding and biological activities. Understanding the structural basis of TCR recognition of HIV epitopes may facilitate the design of vaccines capable of inducing CTLs that recognize HIV epitopes with a higher avidity and that are more effective in preventing HIV infection. We have recently reported that 188Re-linked HIV envelope-specific human mAb can be used to efficiently eliminate HIV-1-infected cells in vivo. We will determine if this approach could be used to develop a new HIV therapy using soluble TCR tetramers coupled to radioisotopes, either in the native form or carrying mutations we identified designed to increase avidity, and examine their in vitro and in vivo capacity to target and eliminate HIV-infected cells. PUBLIC HEALTH RELEVANCE: We propose to perform structure-function analysis of the TCR of HIV-specific CTLs by solving the X-ray crystallographic structures of the complexes formed by HIV-SL9- specific TCRs and cognate MHC Class I:SL9-peptide assemblies. We will define the physical and chemical determinants responsible for MHC:peptide recognition, determine/rationalize the effect of targeted mutations on CTL function and use this information as the basis for the structure-assisted design of novel HIV therapeutics derived from TCRs linked to radionucleotides.